Mammary gland development is essential for the viability of most mammals and the sustainability of many human populations. In addition, exogenous and intrinsic factors that control mammary morphogenesis have repeatedly been implicated in regulating breast cancer risk, tumor subtype, and/or metastatic progression. Thus, identifying the networks that control mammary morphogenesis is necessary for sustaining human development and discovering additional contributors to breast disease. We have recently identified a novel extended family with a breast developmental defect. Half of all males and females in this family have congenital nipple dysmorphia and females fail to develop breasts. The trait follows an autosomal dominant inheritance pattern, does not correspond to any previously described breast developmental defect, and has been mapped to a single chromosomal region that does not contain any established breast development genes. Thus, this family provides a unique opportunity to discover a novel mutation/gene that controls breast development and potentially cancer. With this proposal, deep sequencing will be performed on affected and unaffected family members to identify the specific mutation in this family that segregates with the amastia phenotype. The tissue specific expression of this gene will then be assessed throughout normal mammary gland development in the mouse and in adult human breast tissue. Lastly, we will develop a novel genetically- manipulated mouse model wherein the human mutation has been engineered in the mouse genome. These mice will be a vital resource for establishing the function of the identified gene in breast development and the impact of the human mutation on that function. Upon completion, this R21 proposal will uncover a completely new pathway controlling breast morphogenesis and generate necessary tools for future analyses of the mechanism(s) by which it controls development and possibly cancer.